Optic chiasm involvement in multiple sclerosis, aquaporin-4 antibody-positive neuromyelitis optica spectrum disorder and myelin oligodendrocyte glycoprotein–associated disease

Background: Optic neuritis (ON) is a common feature of inflammatory demyelinating diseases (IDDs) such as multiple sclerosis (MS), aquaporin 4-antibody neuromyelitis optica spectrum disorder (AQP4 + NMOSD) and myelin oligodendrocyte glycoprotein antibody–associated disease (MOGAD). However, the involvement of the optic chiasm (OC) in IDD has not been fully investigated. Aims: To examine OC differences in non-acute IDD patients with (ON+) and without ON (ON−) using magnetisation transfer ratio (MTR), to compare differences between MS, AQP4 + NMOSD and MOGAD and understand their associations with other neuro-ophthalmological markers. Methods: Twenty-eight relapsing-remitting multiple sclerosis (RRMS), 24 AQP4 + NMOSD, 28 MOGAD patients and 32 healthy controls (HCs) underwent clinical evaluation, MRI and optical coherence tomography (OCT) scan. Multivariable linear regression models were applied. Results: ON + IDD patients showed lower OC MTR than HCs (28.87 ± 4.58 vs 31.65 ± 4.93; p = 0.004). When compared with HCs, lower OC MTR was found in ON + AQP4 + NMOSD (28.55 ± 4.18 vs 31.65 ± 4.93; p = 0.020) and MOGAD (28.73 ± 4.99 vs 31.65 ± 4.93; p = 0.007) and in ON− AQP4 + NMOSD (28.37 ± 7.27 vs 31.65 ± 4.93; p = 0.035). ON+ RRMS had lower MTR than ON− RRMS (28.87 ± 4.58 vs 30.99 ± 4.76; p = 0.038). Lower OC MTR was associated with higher number of ON (regression coefficient (RC) = −1.15, 95% confidence interval (CI) = −1.819 to −0.490, p = 0.001), worse visual acuity (RC = −0.026, 95% CI = −0.041 to −0.011, p = 0.001) and lower peripapillary retinal nerve fibre layer (pRNFL) thickness (RC = 1.129, 95% CI = 0.199 to 2.059, p = 0.018) when considering the whole IDD group. Conclusion: OC microstructural damage indicates prior ON in IDD and is linked to reduced vision and thinner pRNFL.


Introduction
Inflammatory demyelinating diseases (IDDs) represent a spectrum of heterogeneous disorders affecting the central nervous system.3][4][5] In MS, ON is often unilateral, characterised by short lesions and tends to recover well. 6,7In contrast, AQP4 + NMOSD-associated ON can be unilateral or bilateral, more frequently associated with severe visual loss and limited recovery if untreated.AQP4 + NMOSD optic nerve lesions are extensive, involving more than half of its length and posteriorly located.In MOGAD, ON is more frequently bilateral and is associated with severe visual loss, yet there is potential for favourable clinical recovery. 8Optic nerve lesions in MOGAD patients are often long and anteriorly located, leading to the frequent observation of optic disc oedema in acute ON.In addition, perineural enhancement has been documented in some cases. 6,9gnetisation transfer (MT) is an advanced magnetic resonance imaging (MRI) technique used to assess the exchange of proton magnetisation between tissue macromolecules and mobile water molecules, a phenomenon typically quantified by the magnetisation transfer ratio (MTR). 10While MTR has been associated with myelin content, MTR changes may also reflect neuroaxonal loss. 11In the optic nerve, MTR has proven to be a valuable measure of early demyelination and predictor of axonal loss and remyelination after acute ON. 12,13 We recently reported that MTR values can also help the discrimination between relapsing-remitting multiple sclerosis (RRMS) and AQP4 + NMOSD. 147][18] These chiasmatic alterations can be related to visual impairment. 9,19However, the comprehensive quantification of OC involvement and its impact on visual outcomes in the context of these three IDDs has not been fully explored.We conducted a prospective study on a cohort of non-acute IDD patients both with (ON+) and without (ON−) previous ON to assess (1) microstructural OC MTR changes between ON+ and ON− patients; (2) whether the degree of the OC MTR changes differs between RRMS, AQP4 + NMOSD and MOGAD and (3) and if OC MTR is associated with residual visual/ophthalmic outcomes in these patients.

Patients and methods
From a previously described cohort, 14 80 patients and 32 healthy controls (HCs) were selected.
Inclusion criteria for patients were (1) diagnosis of RRMS according to 2017 revised criteria 20 or AQP4 + NMOSD according to 2015 Wingerchuk's criteria 21 or MOGAD, defined as MOG-Ab positivity in the context of an acute demyelinating event in patients presenting with a MOGAD phenotype previously described; 22 (2) no clinical relapses in the previous 6 months; (3) no ophthalmic conditions; (4) age above 18 years at the time of assessment and (5) no major contraindications to MRI.The study was conducted in accordance with the International Conference on Harmonisation guidelines for Good Clinical Practice and the Declaration of Helsinki.All participants gave informed consent upon admission to the study, which was approved by the National Research Ethics Service (NRES) Committee London Bloomsbury.
All patients underwent detailed clinical evaluation, peripapillary retinal nerve fibre layer (pRNFL) and macular ganglion cell-inner plexiform layer (GCIPL) thickness measurements with optical coherence tomography (OCT) scanning and MRI.Episodes of ON were identified by clinicians through clinical information collected from medical records.ON was defined as subacute, monocular visual loss associated with pain during eye movement, with objective evidence of an optic neuropathy (e.g.impaired best-corrected visual acuity, dyschromatopsia, relative afferent pupillary defect and/or optic disc pallor/ swelling). 2,23The number of separate inflammatory events was determined for each eye of each patient.Visual assessment was performed for each eye separately with high contrast letter visual acuity using the retro-illuminated Early Treatment Diabetic Retinopathy Study chart at 4 m with best correction.Higher logMAR scores reflect worse visual acuity; a score of 1.7 was assigned when no letters could be correctly identified by the patient.To test the association between MTR value in the OC and the clinical outcome, both (1) visual acuity as an average between the two eyes and (2) visual acuity in the worst eye were entered in separate analyses.
Patients and controls underwent pRNFL and GCIPL OCT scanning using Heidelberg Eye Explore 1.10.2.0 (Spectralis version 6.9a, Heidelberg Engineering, Heidelberg, Germany).Optic nerve thicknesses at 3.4 mm ring scan were extracted.A quality check was performed according to the international OSCAR-IB criteria. 24l participants underwent MRI using a 3T Achieva system (Philips Medical Systems, Best, The Netherlands) and 32-channel head coil based at the Institute of Research, Vall d'Hebron Barcelona Hospital Campus, Barcelona, Spain NMR research unit, Queen Square, London.Left and right optic nerves were acquired separately with (1) coronal-oblique two-dimensional (2D) fat-suppressed turbo spin-echo T2-weighted (T2w) imaging with coronal-oblique plane orthogonal to the long axis of the optic nerve (number of slices 20, slice thickness 3 mm, no slice gap, field-of-view (FOV) 160 × 160 × 160 mm 3 , voxel size 0.5 × 0.5 mm 2 ) and (2) Magnetisation Transfer imaging (MTI), using identical scan geometry as above (number of slices 20, FOV 160 × 160 × 60 mm 3 , acquisition voxel size 0.75 × 0.75 × 3 mm 3 , reconstruction voxel 0.5 × 0.5 × 3 mm 3 ).MTI comprises of a three-dimensional (3D) slab-selective fast field-echo sequence with two echoes, performed with and without Sinc-Gaussian-shaped MT saturating pulses (MTon and MToff, respectively) of nominal angle α = 360°, offset frequency 1 kHz, duration 16 ms.Participants were asked to close their eyes during scanning.Two sets of MT images were acquired for each eye to improve signal-to-noise and were pre-processed to generate MTon average and MToff average and then co-registered to native T2w space.
The average number of slices for the OC was 1.73 slices (median = 1, range = 1-2).Chiasmal MTR processing is described in Figure 1.Manually delineated Regions of Interest (ROIs) were created slice-by-slice to encompass the right and left sections of the OC (optic hemichiasms) independently on each T2w acquisition.OC ROIs were delineated on the T2w coronal orbital MRI scans by two experienced raters (R.C. and A.B.) for left and right optic nerve acquisitions using JIM 6.0 (Xinapse systems, http://www.xinapse.com).Raters were blinded to diagnosis during region of interest (ROI) delineation.There was a high degree of consistency in interrater agreement in identifying the optic nerve ROIs, as indicated by a 95% Cohen's kappa coefficient.ROIs were subsequently transferred from T2w images to the pre-registered MTon and MToff average images and manually adjusted to ensure were manually delineated slice-by-slice to cover the right and left section of the optic chiasm independently on each T2w acquisition.Afterwards, these ROIs were transposed onto the pre-registered MTon and MToff average images and manually adapted to delineate the same area on the pre-registered MTon and MToff average.Subsequently, to correct for small nerve motion, the three ROIs' centre-of-mass slice-wise were aligned to a common space (T2w and registered average MTon and MToff) and (c) MTR values for both left and right optic hemichiasms were obtained from the registered images.For each patient, right and left MTR values were averaged to obtain a single averaged chiasmal MTR value that was then impute into the statistical analysis.consistent delineation of the same area on T2w, MTon and MToff images.To additionally account for minor nerve motion, the centre-of-mass of the three ROIs was aligned slice-wise to a common space built on T2w and registered average MTon and MToff images.The registered images were then utilised to extract MTR values for both the left and right optic hemichiasms.Finally, the MTR values from the hemichiasms were statistically averaged to obtain a single chiasmal MTR value.Clinical and demographic characteristics of groups were compared using t-tests or analysis of variance (ANOVA) for continuous variables and Chi-square test for independence to compare the groups in categorical variables.

Statistical analysis
Multivariable linear regression models were fitted to estimate the associations between OC MTR as the continuous response variable and different clinical categorical predictors described above, 25 adjusting for potential confounders, including age, sex, disease duration and MRI scanner software upgrade which occurred during the study.
We also tested associations between (1) OC MTR versus visual acuity and (2) OC MTR versus pRNFL and GCIPL independently.Linear regression models were fitted with logMAR acuity, pRNFL thickness or GCIPL thickness as outcomes in two ways: (1) average logMAR and average pRNFL/GCIPL across both eyes or (2) worse logMAR and thinner pRNFL/ GCIPL between both eyes for each individual.OC MTR and other potential confounders were entered as independent variables.
All inferences used a type I error rate of p < 0.05 for clear statistical significance.
IDD patients had average disease duration of 8.2 ± 7.1 years.Disease duration did not differ significantly between ON+ IDD and ON− IDD.Fortyeight patients (60%) experienced at least one previous ON, and among them, ON affected both eyes in 21/48 (43.75%) and was relapsing in 21/48 (39.58%).The average number of inflammatory events was 2.52 ± 2.32, with 22/48 (45.83%) patients reporting a single episode, 10/48 (20.83%) having experienced two previous ON and 16/48 (33.33%) reporting three or more previous ON.The average time-gap from the first ON was 87.5 ± 76.2 months, while the time-gap from the last one was 51.1 ± 52.5 months.Clinical characteristics of the different disease groups are summarised in Table 1.The prevalence of previous ON and the mean number of ON were higher in MOGAD and AQP4 + NMOSD when compared with RRMS (both p < 0.001).The same groups reported higher percentages of bilateral ON (p = 0.011) and relapsing ON episodes (p = 0.002).

Effect of previous ON on OC MTR in IDD
Chiasmal MTR values in ON− and ON+ patients are reported in Table 3. ON+ IDD patients showed lower MTR values compared with HCs (regression coefficient (RC) = −3.33,95% confidence interval (CI) = −5.56 to −1.111, p = 0.004) and slightly lower MTR values compared with ON− IDD patients although non-significant (Figure 2).OC MTR values were not significantly different between HCs and ON− IDD patients.

Discussion
This study provides insights into the mechanisms of OC damage in IDD with and without previous ON.ON+ IDD patients showed lower MTR values compared with HCs and ON− IDD patients, with a higher number of previous ON episodes associated with lower OC MTR.
In our study population, chiasmal involvement was more pronounced in patients with AQP4 + NMOSD and MOGAD than in those with MS.In addition, lower OC MTR values correlated with reduced pRNFL thickness and poorer visual outcomes.
The strong association between the number of previous ON episodes and chiasmal MTR aligns with previous research on post-acute changes following inflammatory optic nerve lesions. 17,26Lower MTR values are likely to reflect reduced microstructural integrity 10 which can result from both demyelination and inflammatory-related changes during the acute phase of ON, 13,27 while correlate with demyelination, remyelination and neuroaxonal loss in the longterm, 28-30 representing a useful marker of chronic damage and repair. 13 16 However, these processes may be incomplete and the occurrence of new acute ON episodes could progressively exhaust the mechanisms responsible for recovery.Therefore, we hypothesise that the greater reduction in OC MTR following multiple ON episodes could result from two related factors: direct axonal transection and degeneration caused by multiple inflammatory attacks and/or a progressive impairment of the post-inflammatory recovery processes due to the accumulation of damage associated with relapsing ON.This hypothesis gains support from the correlation we observed between OC MTR values and pRNFL thickness, a well-recognised marker of neuroaxonal loss in the anterior visual pathways.2][33] The association we observed between OC MTR values and pRNFL in our study underscores the role of MTR as a measure of chronic biological damage in the anterior optic pathway.Interestingly, we did not observe significant relationships between GCIPL and chiasmal metrics.However, GCIPL data were available for only 49 out of 80 patients.It is likely, therefore, that this analysis was underpowered to be able to establish significance, possibly as a consequence of high standard errors even though the association was in a physiologically plausible direction (RC = 0.554, 95% CI = −0.371 to 1.480, p = 0.235).
Our study also revealed an association between lower OC MTR values and poorer visual acuity.The OC is recognised as a region with high axonal density, and its damage has been linked to a decline in visual function. 19Our results suggest that microstructural damage occurring in both the optic nerve and OC following ON indicates a broader pattern of microstructural damage.This damage appears to contribute to a reduction in overall visual function, in line with previous studies. 9,19The linear association we observed between OC MTR measures and visual acuity further emphasises the role of chiasmal MTR as a useful measure for assessing both biological and clinical damage in the anterior visual pathway.
Of note, we observed that in MOGAD, there was no association between visual acuity and MTR measures.This finding is consistent with the observation that, despite significant microstructural damage of the anterior visual pathway, MOGAD patients exhibit better functional recovery compared with AQP4 + NMOSD. 7,8Further studies will be necessary to better understand the mechanisms underlying this function-structure discordance.
The involvement of the OC in ON lesions has been well documented in cases of AQP4 + NMOSD, but it is less common in MS and MOGAD. 2,15Despite these differences, in our population we observed that ON+ AQP4 + NMOSD and MOGAD had lower MTR values when compared to HCs.In contrast, ON+ RRMS showed MTR values that were non-significantly lower than those of the control group.These findings align with recent research which challenges the conventional assumption that the frequency of OC involvement in AQP4 + NMOSD and MOGAD significantly differs. 15While the percentage of OC involvement in these two antibody-mediated diseases is not dissimilar (20% in AQP4 + NMOSD vs 16% in MOGAD), Tajfirouz et al. 15 noted that the characteristics of OC damage seem to vary between these conditions.Specifically, the authors proposed that OC involvement in MOGAD could be attributed to an 'extension' of long anterior lesions, whereas in AQP4 + NMOSD, the damage may be more directly associated with the posterior localisation of the lesion. 15Interestingly, in both disorders, the damage appears to be secondary to an inflammatory process within the optic nerve.Our findings support this hypothesis, by revealing that the higher number of ON episodes recorded in MOGAD and AQP4 + NMOSD patients (with a mean of 2.93 and 1.13, respectively) resulted in more severe damage of the OC, as indicated by lower MTR values.However, we also observed that in MOGAD and AQP4 + NMOSD patients, OC MTR values remained lower than those of HCs, even after accounting for the number of previous ON episodes.This observation might suggest (1) the existence of damage independent from relapse which could subtly contribute to the decline in microstructural integrity within the OC and/or (2) the occurrence of subclinical ON.This phenomenon is notably pronounced in AQP4 + NMOSD, who reported lower MTR values than those of RRMS patients.In addition, in this group, ON− patients showed lower MTR values compared with HCs.Juenger et al. 19 have previously reported a reduction in chiasmal dimensions in ON− NMOSD, suggesting that microstructural changes could occur in the optic pathway of NMOSD patients independent of previous ON.Our results corroborate this hypothesis, underscoring the complex relationship between ON, OC involvement and the distinct characteristics of IDD subtypes and emphasising the importance of considering these factors when assessing the impact of these diseases on visual function and structural integrity.
Our study has limitations: first, it adopts a cross-sectional design, and we acknowledge that a longitudinal, prospective cohort approach would better assess damage and repair processes during the post-acute phase.Second, the absence of images acquired for the evaluation of optic nerve lesions during the acute phase limited our ability to assess the influence of these lesions on chronic optic nerve damage.However, we addressed this second limitation by building robust statistical models and creating associations based on the available data, including OCT and visual data and volume of brain white matter lesions.Finally, we recognise that the OC is a small structure, as is the optic nerve, making investigation of the anterior visual pathway challenging.Nonetheless, compared with the optic nerve, the OC is less susceptible to motion artefact, less variable in morphology and possesses larger in dimensions, reinforcing its viability as a target for quantitative MRI.In addition, the fact that our results, demonstrate significant associations between previous optic neuritis and OC damage, along with strong associations between OCT and clinical data, support the proposition that this structure could play an important role as an accessible target for assessing the visual pathway in inflammatory diseases.
In conclusion, our findings suggest that OC damage in ON, as demonstrated by MTR, is linked to the number of inflammatory ON episodes experienced by IDD patients and reflects pathological changes occurring in the anterior visual pathways within IDD.As optic nerve MRI remains challenging due to its small dimensions, mobility and susceptibility to artefacts, MTR OC imaging could provide additional information for evaluating the anterior visual pathways in IDDs.

Figure 1 .
Figure 1.The figure illustrates the processing pipeline for the magnetisation transfer ratio (MTR) optic pathway analysis: (a) Two MTR acquisitions (2 echo-times) were obtained for each eye and they were pre-processed to generate two MTon and two MToff images.From these two different echo time MT images, average MToff and average MTon images were created, which were subsequently pre-registered to the native T2w image, (b) regions of interest (ROIs) were manually delineated slice-by-slice to cover the right and left section of the optic chiasm independently on each T2w acquisition.Afterwards, these ROIs were transposed onto the pre-registered MTon and MToff average images and manually adapted to delineate the same area on the pre-registered MTon and MToff average.Subsequently, to correct for small nerve motion, the three ROIs' centre-of-mass slice-wise were aligned to a common space (T2w and registered average MTon and MToff) and (c) MTR values for both left and right optic hemichiasms were obtained from the registered images.For each patient, right and left MTR values were averaged to obtain a single averaged chiasmal MTR value that was then impute into the statistical analysis.

Figure 3 .
Figure 3. Association between magnetisation transfer ratio (MTR) values and number of previous optic neuritis (ON) episodes.Predicted chiasmal MTR values from number of previous optic neuritis from the linear regression model adjusted for age, sex and MRI upgrade.

Figure 4 .
Figure 4. Comparison between magnetisation transfer ratio (MTR) values in the optic chiasm (OC) and number of previous optic neuritis (ON) episodes in inflammatory demyelinating diseases (IDD).

Figure 6 .
Figure 6.Association between peripapillary retinal nerve fibre layer (pRNFL) thickness and magnetisation transfer ratio (MTR) values in the optic chiasm (OC).Visual acuity is reported as average pRNFL thickness between the two eyes.Adjusted predictions of visual acuity from chiasmal MTR values from the linear regression model adjusted for age, sex and MRI upgrade.

Figure 7 .
Figure 7. Association between visual acuity (VA) and magnetisation transfer ratio (MTR) values in the optic chiasm (OC).VA is reported as average visual acuity between the two eyes.Adjusted predictions of visual acuity from chiasmal MTR values from the linear regression model adjusted for age, sex and MRI upgrade.

Table 1 .
Clinical characteristics of the population of study.
RRMS: relapsing-remitting multiple sclerosis; AQP4 + NMOSD: aquaporin 4-antibody neuromyelitis optica spectrum disorder; MOGAD: myelin oligodendrocyte glycoprotein antibody-associated disease; HCs: healthy controls.a p-value using ANOVA (Analysis of Variance) test to compare the groups.b Chi-square test for independence to compare the groups.

Table 2 .
Clinical characteristics of the population of study.
ON+ IDD: patients with inflammatory demyelinating diseases and at least a previous optic neuritis episode; ON− IDD: patients with inflammatory demyelinating diseases without history of previous optic neuritis episode; HCs: healthy controls.a p-value using ANOVA (Analysis of Variance) test to compare the groups.b Chi-square test for independence to compare the group.